去极化反跳调控脊髓背角神经元兴奋性的机制研究

Chronic pain is highly prevalent and our understanding of its underlying mechanisms is incomprehensive. It seriously impacts on patients’ well-being as well as life quality, and its clinical management is still a huge challenge. Rebound depolarization is related to T-type calcium current (IT) and hyperpolarization–activated current (Ih). It is a subsequent response to hyperpolarizing stimulation and plays a critical role in the development and maintenance of chronic pain. Spinal substantia gelatinosa (SG) neurons have been identified as the “central gate” of transmitting and modulating nociceptive information. However, the morphological and electrophysiological features, together with the synaptic connections of neurons exhibiting rebound depolarization remain unclarified. Our preliminary studies found that inhibition of Ih decreases the frequency of action potential and depresses the excitability of SG neurons. Therefore, we hypothesis that rebound depolarization could modulate local pain signaling by altering neuronal excitability in SG. By using combination of whole-cell patch-clamp, immunohistochemistry, and single-cell RT-PCR methods, we will elucidate the morphological and electrophysiological properties as well as physiological function of neurons exhibiting rebound depolarization. We will further illustrate the morphological and electrophysiological properties of neurons which pre- and post- connect to those rebound depolarizing neurons. Together, this study will interpret a new mechanism underlying chronic pain and suggest potential targets for clinical treatment of chronic pain.

慢性疼痛发病率高，发病机制不清，严重影响患者的身心健康和生活质量，临床防治任重道远。去极化反跳是部分神经元接受超极化刺激后的固有反应，可能与T型钙电流（IT）和超极化激活阳离子电流（Ih）相关，在慢性疼痛的发生发展中具有关键作用。脊髓背角胶状质（SG）神经元是痛觉的传递与整合的“闸门”，SG表达去极化反跳神经元的形态、电生理特性及其突触联系不明。我们的前期研究发现，抑制SG的Ih可降低动作电位发放频率、减弱神经元兴奋性。我们推测，去极化反跳可通过调控SG神经元兴奋性，参与疼痛局部神经回路的调制。本项目拟结合全细胞膜片钳、免疫组织化学、单细胞RT-PCR等方法，揭示大鼠SG去极化反跳神经元及其突触前、后神经元的形态、电生理特性，进一步研究SG神经元去极化反跳的特征与功能，从而完善慢性疼痛的脊髓机制，为慢性疼痛临床治疗提供新靶点及理论依据。

去极化反跳是指神经元受到超极化脉冲刺激后随即出现的短暂膜电位去极化反应，其产生可能与超极化环核苷酸门控阳离子通道、T型钙通道密切相关。该现象作为神经元的基本膜特性已在丘脑、小脑等不同脑区的神经元上记录到。但在伤害性信息上行传递的中枢门户，即脊髓背角胶状质区该现象的特点和形成机制如何？SG区具有去极化反跳的神经元是否具有电生理和/或形态学特异性？SG区去极化反跳在神经病理性疼痛状态下有何变化？我们的研究发现：1、与无去极化反跳现象的SG神经元相比，具有去极化反跳现象的神经元RMP更正、基电流更小、AP阈值更负、AP半宽度更短，提示去极化反跳神经元固有兴奋性更高；2、具有去极化反跳现象的SG神经元放电模式以tonic为主、形态分类以islet为主，说明去极化反跳现象在脊髓背角SG区主要存在于抑制性中间神经元；3、HCN通道介导的Ih以及T型钙通道介导的IT是大鼠脊髓背角去极化反跳形成的关键离子基础：Ih决定去极化反跳首发放电潜伏期，IT影响去极化反跳振幅；4、作为去极化反跳现象形成的重要离子基础，IT以及介导这一电流的T型钙通道（主要为Cav3.2亚型）在PSNL所致神经病理性疼痛模型中其重要作用，抑制该通道电流具有良好的镇痛作用；5、PSNL过程中脊髓背角HCN2这一亚型的mRNA及蛋白表达水平均显著上调；6、病理性疼痛状态下，脊髓背角兴奋性中间神经元兴奋性上调，表现为非tonic放电的AP阈值更负、AP阈值与RMP的差值减小；抑制性中间神经元兴奋性下调，表现为去极化反跳现象中spike放电频率减低。本项目通过揭示SG神经元去极化反跳的特征与功能，为临床神经病理性疼痛的防治提供了新思路和理论依据。